Various types of polysaccharides are known, such as plant-derived polysaccharides (e.g., starch and cellulose), microorganism-derived polysaccharides (e.g., xanthan), and higher organism-derived polysaccharides (e.g., hyaluronan, heparin, and chondroitin). These polysaccharides are used in medicinal products, food products, general industrial applications, and other various applications.
Hyaluronan, for example, is found in high quantity in biological tissues, such as bovine eyes, rooster combs, shock-absorbing tissue of animals, placentas, cancer cells, and skin. Hyaluronan is a linear polysaccharide consisting of alternating glucuronic acid and N-acetylglucosamine bonded by β1,3 and β1,4 linkages, and is a high molecular weight glucosaminoglycan having a molecular weight of 105 to 106 Da. Hyaluronan is characterized by its high viscosity, high moisturizing effect, excellent lubricating effect against physical friction, and protective effect against bacterial and other invasions.
Because of these characteristics, hyaluronan is widely used as a cosmetic additive and a pharmaceutical additive (e.g., arthritis treatment agent, wound dressing agent, ophthalmic surgery adjuvant, and post-surgery anti-adhesion agent).
Widely known methods for producing hyaluronan include: (1) a method for extracting hyaluronan from the biological tissues (extraction method) (Patent Literatures 1 and 2), and (2) a method for collecting hyaluronan from a culture of microorganisms having an ability to produce hyaluronan in the presence of a sugar such as glucose (microorganism culture method) (Patent Literatures 3 and 4).
Unfortunately, the hyaluronan produced by the extraction method (1) contains impurities such as chondroitin sulfate and glycosaminoglycan sulfate. Hence, a complicated purification process is required in order to remove these impurities.
In the hyaluronan production by the microorganism culture method (2), an increase in the hyaluronan production entails an increase in the viscosity of the culture medium, making it difficult to stir for aeration. As stirring for aeration becomes more difficult, the hyaluronan production will eventually stop. Thus, the microorganism culture method unfortunately has very low hyaluronan production efficiency. In addition, because of the high viscosity of the culture medium, the method requires a complicated purification process in order to remove the microorganisms used. Further, hyaluronan-degrading enzymes present in the microorganisms degrade the produced hyaluronan, causing problems such as no increase in the molecular weight of the hyaluronan and high heterogeneity in the molecular weight.
Also in the case of other polysaccharides besides hyaluronan, the extraction method involves contamination with various impurities, and the microorganism culture method has drawbacks such as low production efficiency, no increase in the molecular weight, and high heterogeneity in the molecular weight.
Thus, a method for producing a polysaccharide using a polysaccharide synthase (enzymatic synthesis method) has been considered as a third method that does not involve biological tissue extraction or microorganism cultivation. For example, a method that uses a hyaluronan synthase is known (Non-Patent Literature 1). Unfortunately, the enzymatic synthesis method has drawbacks such as requirement for a large amount of enzyme, low production efficiency, and a low yield. Thus, the method remains at the laboratory level and its application at the industrial level is yet to be considered.